Project Summary Aging-related alterations of endothelial cells play a central role in the pathogenesis of many chronic human diseases, including but not limited to cardiovascular diseases, sarcopenia, Alzheimer's disease, and tumor progress/metastasis. By elucidating the molecular mechanisms underlying aging-related endothelial dysfunction, new pathways could be revealed for designing novel diagnostic biomarkers and identifying possible therapeutic targets for those common and devastating diseases. Recent evidence from different types of studies ? gene microarray, epidemiologic, and cultured tumor endothelial cells ? strongly indicates that cardiac troponin T (cTnT) is expressed in vascular endothelial cells. cTnT is a known A kinase anchoring protein. We reported recently that it regulates the protein kinase A (PKA) regulatory subunit RI and RII differentially at neuromuscular junction in skeletal muscle of old mouse, indicating and age and tissue type- dependent PKA regulation by cTnT. A kinase anchoring proteins also interact with protein kinase C (PKC). Both PKA and PKC are involved in caspase activation, and these molecules play critical roles in regulating endothelial barrier integrity. The latter is critical for the maintenance of water and protein balance between the intravascular and extravascular compartments. An impairment of endothelial barrier integrity and function (for instance, increased permeability of blood-brain barrier) has been implicated in the genesis and/or progression of a variety of pathological conditions and diseases, including Alzheimer's disease. Our preliminary data revealed that cTnT expression in vascular endothelial cells is higher in old mice than the young, and is associated with higher levels of caspase-3 activation. Given all these evidence, our central hypotheses are that (a) cTnT expression in vascular endothelial cells increases with aging, and (b) this change leads to impaired endothelial barrier integrity through altering PKA/PKC-caspase signaling. In this exploratory R21 application, led by a New Investigator, we propose 3 Specific Aims to test our hypothesis. Aim 1 seeks to determine if age-dependent change in cTnT is associated with altered PKA, PKC, and caspases in brain vascular endothelial cells. Aim 2 seeks to determine if cTnT-PKA/PKC-caspase signaling regulates vascular endothelial cell death. Aim 3 seeks to determine if manipulation of cTnT expression level in brain vascular endothelial cells in vivo can affect integrity of the blood-brain barrier in old mice. This project, conducted by a multidisciplinary team, will be the first to define a role for cTnT in vascular endothelial cell aging. Findings may help provide insights into new molecular mechanisms and possible therapeutic options for Alzheimer's disease and other aging-related chronic diseases.